Binding Thermodynamics of Phosphorylated Inhibitors to Triosephosphate Isomerase and the Contribution of Electrostatic Interactions

Serratos, Iris N.; Pérez-Hernández, Gerardo; Garza‐Ramos, Georgina; Hernández‐Arana, Andrés; González-Mondragón, Edith; Zubillaga, Rafael A.

doi:10.1016/j.jmb.2010.10.018

Cited by 8 publications

(5 citation statements)

References 61 publications

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“…A 0.1 μM VC1 solution (2 mL) was titrated with successive volumes of 0.02 μM and 0.2 μM QUIN. We adjusted the titration data set according to previous authors [48,49], assuming that the fluorescence intensities of the emitting species (free VC1 and VC1–QUIN complex) are additive, according to Equation 1, where x is the total concentration of inhibitor in the cell, Et represents the total concentration of VC1, K d is the dissociation equilibrium constant ( K d = 1/ K b ) and a denotes the asymptotic value to which Y tends at high x values. Y = 1-F/F t , were F is the overall fluorescence intensity after each addition of QUIN and F t is the fluorescence of free VC1 at the corresponding concentration, respectively.…”

Section: Methodsmentioning

confidence: 99%

Modeling the Interaction between Quinolinate and the Receptor for Advanced Glycation End Products (RAGE): Relevance for Early Neuropathological Processes

et al. 2015

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The receptor for advanced glycation end products (RAGE) is a pattern-recognition receptor involved in neurodegenerative and inflammatory disorders. RAGE induces cellular signaling upon binding to a variety of ligands. Evidence suggests that RAGE up-regulation is involved in quinolinate (QUIN)-induced toxicity. We investigated the QUIN-induced toxic events associated with early noxious responses, which might be linked to signaling cascades leading to cell death. The extent of early cellular damage caused by this receptor in the rat striatum was characterized by image processing methods. To document the direct interaction between QUIN and RAGE, we determined the binding constant (Kb) of RAGE (VC1 domain) with QUIN through a fluorescence assay. We modeled possible binding sites of QUIN to the VC1 domain for both rat and human RAGE. QUIN was found to bind at multiple sites to the VC1 dimer, each leading to particular mechanistic scenarios for the signaling evoked by QUIN binding, some of which directly alter RAGE oligomerization. This work contributes to the understanding of the phenomenon of RAGE-QUIN recognition, leading to the modulation of RAGE function.

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Section: Methodsmentioning

confidence: 99%

Modeling the Interaction between Quinolinate and the Receptor for Advanced Glycation End Products (RAGE): Relevance for Early Neuropathological Processes

et al. 2015

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“…The experimentally obtained enthalpy of unfolding for the ScTIM complex is 230 kcal·mol −1 and increases to 284 kcal·mol −1 when PGH is bound. The difference (54 kcal·mol −1 ) is in excellent agreement with the experimentally obtained binding enthalpy (53.7 kcal·mol −1 ) . In contrast, Δ H obs for the TIM63‐PGH complex is 276 kcal·mol −1 , 60% higher than that of the empty form.…”

Section: Resultssupporting

confidence: 85%

“…the non‐catalytic side of the barrel (see below). Throughout this work, the TIM from Saccharomyces cerevisiae (ScTIM) was selected as a reference for comparison, because its function, structure, folding, stability and catalytic mechanism have been thoroughly characterized . All ancTIMs selected for experimental characterization possess the conserved catalytic triad (K12, H95 and E165 in ScTIM, corresponding to K14, H107 and E182 in the multiple alignment shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The estimated ligand‐binding enthalpy Δ H b at the T 1/2 of TIM63 is 57 kcal·mol −1 (estimated from the Δ H b at 25 °C shown in Fig. and the ΔCp b reported in ). Therefore, in the presence of PGH, Δ H tot should be 219 kcal·mol −1 , a value similar to Δ H tot obtained for extant TIMs in the absence of PGH (210–266 kcal·mol −1 ) , and 47 kcal·mol −1 higher than Δ H tot obtained in the absence of PGH (172 kcal·mol −1 ).…”

Section: Resultsmentioning

confidence: 99%

“…M: monomer; M 2 : dimer; L: ligand. The estimation of Δ H b at different temperatures was done with the experimentally obtained Δ H b at 25 °C and the ΔCp b reported in .…”

Section: Methodsmentioning

confidence: 99%

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Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

et al. 2019

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Function, structure, and stability are strongly coupled in obligated oligomers, such as triosephosphate isomerase (TIM). However, little is known about how this coupling evolved. To address this question, five ancestral TIMs (ancTIMs) in the opisthokont lineage were inferred. The encoded proteins were purified and characterized, and spectroscopic and hydrodynamic analysis indicated that all are folded dimers. The catalytic efficiency of ancTIMs is very high and all dissociate into inactive and partially unfolded monomers. The placement of catalytic residues in the three‐dimensional structure, as well as the enthalpy‐driven binding signature of the oldest ancestor (TIM63) resemble extant TIMs. Although TIM63 dimers dissociate more readily than do extant TIMs, calorimetric data show that the free ancestral subunits are folded to a greater extent than their extant counterparts are, suggesting that full catalytic proficiency was established in the dimer before the stability of the isolated monomer eroded. Notably, the low association energy in ancTIMs is compensated for by a high activation barrier, and by a significant shift in the dimer‐monomer equilibrium induced by ligand binding. Our results indicate that before the animal and fungi lineages diverged, TIM was an obligated oligomer with substrate binding properties and catalytic efficiency that resemble that of extant TIMs. Therefore, TIM function and association have been strongly coupled at least for the last third of biological evolution on earth. Databases PDB Entry: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=6NEE. Enzymes Triosephosphate isomerase http://www.chem.qmul.ac.uk/iubmb/enzyme/EC5/3/1/1.html, Glycerol‐3‐phosphate dehydrogenase http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/1/1/8.html.

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Stable monomers in the ancestral sequence reconstruction of the last opisthokont common ancestor of dimeric triosephosphate isomerase

Pérez‐Niño,

Guerra,

Díaz‐Salazar

et al. 2024

Protein Science

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Function and structure are strongly coupled in obligated oligomers such as Triosephosphate isomerase (TIM). In animals and fungi, TIM monomers are inactive and unstable. Previously, we used ancestral sequence reconstruction to study TIM evolution and found that before these lineages diverged, the last opisthokonta common ancestor of TIM (LOCATIM) was an obligated oligomer that resembles those of extant TIMs. Notably, calorimetric evidence indicated that ancestral TIM monomers are more structured than extant ones. To further increase confidence about the function, structure, and stability of the LOCATIM, in this work, we applied two different inference methodologies and the worst plausible case scenario for both of them, to infer four sequences of this ancestor and test the robustness of their physicochemical properties. The extensive biophysical characterization of the four reconstructed sequences of LOCATIM showed very similar hydrodynamic and spectroscopic properties, as well as ligand‐binding energetics and catalytic parameters. Their 3D structures were also conserved. Although differences were observed in melting temperature, all LOCATIMs showed reversible urea‐induced unfolding transitions, and for those that reached equilibrium, high conformational stability was estimated (ΔGTot = 40.6–46.2 kcal/mol). The stability of the inactive monomeric intermediates was also high (ΔGunf = 12.6–18.4 kcal/mol), resembling some protozoan TIMs rather than the unstable monomer observed in extant opisthokonts. A comparative analysis of the 3D structure of ancestral and extant TIMs shows a correlation between the higher stability of the ancestral monomers with the presence of several hydrogen bonds located in the “bottom” part of the barrel.

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Binding Thermodynamics of Phosphorylated Inhibitors to Triosephosphate Isomerase and the Contribution of Electrostatic Interactions

Cited by 8 publications

References 61 publications

Modeling the Interaction between Quinolinate and the Receptor for Advanced Glycation End Products (RAGE): Relevance for Early Neuropathological Processes

Modeling the Interaction between Quinolinate and the Receptor for Advanced Glycation End Products (RAGE): Relevance for Early Neuropathological Processes

Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

Stable monomers in the ancestral sequence reconstruction of the last opisthokont common ancestor of dimeric triosephosphate isomerase

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